Throughout this application, various publications are referenced and full citations for these publications may be found in the references at the end of the specifications preceding the claims. The disclosures of these publications are hereby incorporated by reference into this application in order to more fully describe the state of the art as known to the skilled therein as of the date of the invention described and claimed herein.
Fagopyrum cymosum (Trev.) Meisn is a species of buckwheat, which belongs to the Genus Fagopyrum, family Polygonaceae. There are about 14 species of buckwheat, either cultivated or wild, existing around the world. Buckwheat grows mainly in the temperate area of Asia, Europe and North America. (1)
China was the earliest country in the world to cultivate buckwheat. According to archaeological studies, China began to grow buckwheat about 2,000 years ago. The genetic origins of buckwheat are believed to occur in Yunnan and Sichuan Provinces of China.(2) Buckwheat was brought from China to Japan via Korea Peninsula and then to Europe via Siberia and southern Russia. Germany was the first country in Europe to grow buckwheat in 1396. Then the crop was introduced into Belgium, France, Italy and Britain in the 17th century. Later buckwheat was brought to North America from the Netherlands.
At present, there are three best known species of buckwheat: common buckwheat (Fagopyrum esculentum), tartary buckwheat (Fagopyrum tataricum) and cymose buckwheat (Fagopyrum cymosum). The common buckwheat (Fagopyrum esculentum) is widely cultivated in the Northern Hemisphere. In China, it grows mainly in the northern and northwestern provinces. The tartary buckwheat (Fagopyrum tataricum) is cultivated primarily in the Himalayan area and south and southwest China. The cymose buckwheat (Fagopyrum cymosum) is basically a wild species, not available for human consumption as a crop, although some reports indicate that it is also cultivated in China on a small scale. The cymose buckwheat (Fagopyrum cymosum) distributes only in south and southwest China.
Although buckwheat is cultivated mainly as a minor crop around the world, it has been found recently that certain species of buckwheat also possess therapeutic properties. Animal tests and clinical trials in China have indicated that tartary buckwheat flour is effective in preventing and treating diabetes, hypertension, cerebral arteriosclerosis and cardiovascular disease. It also has the function of invigorating the stomach, facilitating digestion, enhancing the immune system and alleviating inflammation.(3)
The most important therapeutic property discovered in buckwheat is the anti-cancer effect of certain buckwheat species. For example, Samel, D. et al. reported in 1996 that they had examined the effect of a purified extract of the flowering herb of Fagopyrum esculentum on various protein kinases involved in signal transduction, finding that Fagopyrum esculentum contains red fluorescent compounds having photosensitizing properties. Spectrophotometric analysis of the extract indicated structural similarity to hypericin. Dose- and light-dependent inhibition of various protein kinases was observed. The purified Fagopyrum esculentum extract strongly inhibited two receptor-associated protein tyrosine kinases (EGF-R and Ins-R) and a Ser/Thr kinase (PK-C) at an ng/ml concentration range. Selectivity was exhibited as a decreased sensitivity to cytosolic PTKs and protein kinase CK-2. The protein kinases are important components of the signal transduction pathway. Aberration of signal transduction is a hallmark of several proliferative diseases. The researchers believed that the results of their experiment indicated that photosensitizing compounds in Fagopyrum esculentum are potential antiproliferative agents.(4)
In China, research on anti-cancer effect of buckwheat species was concentrated on Fagopyrum cymosum (Trev.) Meisn, which is also termed as Fagopyrum dibotrys (D. Don) Hara.
Fagopyrum cymosum (Trev.) Meisn has been used as a herbal medicine in China since ancient times, but mainly in combination with other medicinal herbs. Modern clinical studies conducted in China have shown that preparations of Fagopyrum cymosum (Trev.) Meisn can be used to treat lung abscess, bacillary dysentery and pyogenic infections. (5) Most importantly it has been found that preparations of Fagopyrum cymosum (Trev.) Meisn can be applied effectively for therapeutic purposes to various kinds of tumors.
Liu, W F et al. published an article in 1981 on some pharmacological properties of Fagopyrum cymosum (Trev.) Meisn, the root of which had been used for the treatment of pulmonary abscess. Liu pointed out that major active principle of Fagopyrum cymosum (Trev.) Meisn is flavanol (5, 7, 3xe2x80x2, 4xe2x80x2-tetrahydroxyflan-3-01 dipolymers). Neither the extract of Fagopyrum cymosum (Trev.) Meisn nor flavanol exhibited significant antibacterial action in vitro. No antibacterial substance was found in the urine of either mice or human beings nor was it found in the organs of mice given the extract p. o. Liu indicated that therapeutic effect of Fagopyrum cymosum (Trev.) Meisn was shown in mice infected I. p. with staphylococcus aureus only when the extract of Fagopyrum cymosum (Trev.) Meisn or flavanol was given via the same route. Phagocytic action of peritoneal macrophages was enhanced, but the number of macrophages was not increased when the extract of Fagopyrum cymosum (Trev.) Meisn or flavanol was given I. p. to mice. Antipyretic action was demonstrated in rabbits. Flavanol was shown to be anti-inflammatory in mice and rats. Platelet aggregation in rats induced by ADP and staphylococcus aureus in vitro was inhibited by flavanol. The same effect on platelet aggregation induced by ADP and collagen was found when flavanol was given IV to rats. Flavanol given IV showed also expectorant effect in mice as shown by the phenol red method, but the bronchial excretion was not increased.(6)
Liu, Y L et al. in 1983 reported their research results on the chemical constituents of Fagopyrum cymosum (Trev.) Meisn. Liu indicated that they had isolated three components of Fagopyrum cymosum (Trev.) Meisn. Component A is the main constituent in Fagopyrum cymosum (Trev.) Meisn and accounts for its pronounced therapeutic effect on pulmonary abscess. The octamethylether, octamethylether diacetate and decanacetate derivatives were prepared from this component. On the basis of spectroscopic analyses, degradation products and physico-chemical constants, component A was identified as the dimer of 5, 7, 3xe2x80x2, 4xe2x80x2-tetrahydroxyflavan-3-ol (C4-C8 linked), named dimeric procyanidin. Components B and C were identified as hecogenin and xcex2-sitosterol respectively.(7)
Liu T C published an article in 1983 analyzing the growth and accumulation of active constituents during different development stages of Fagopyrum cymosum rhizome. The author reported that condensed procyanidin, the active constituent of Fagopyrum cymosum rhizome was observed and studied, including its existence in different parts of the plant and at different development stages, and its development in the root, stem and the above-the-ground part in different growth stages. The paper showed that the amount of condensed procyanidin is higher in the root and stem and is highest from mid-October till the withering season. The content rapidly increases from mid-June and reaches its highest level in late October.(8)
Yao R C et al. reported anti-tumor active constituents of cymose buckwheat in China in 1989. The researchers reported that they had extracted the active portion (A) from rhizome of cymose buckwheat using ethanol extraction, macroporous resin (D101) column chromatography and acetone extraction process. Chemical analysis revealed that A is a compound of proanthocyanidin, from which, in comparison with standard substances, (xe2x88x92) epicatechin, 3-galloyl (xe2x88x92) epicatechin, procyanidin B-2 and B-4, and 3,3xe2x80x2-digalloyl procyanidin B-2 had been detected. In the meantime, methylated, acetylated and trimethyl silica etherified derivatives of A had been prepared.(9)
Ma Y P et al. published an article in 1989 in China on prediction of responsiveness of human lung cancer xenograft to extracts of Fagopyrum cymosum (Trev.) Meisn by SRC assay. The researchers reported that they had determined responsiveness of human lung cancer explant to extracts E and CD1 of Fagopyrum cymosum (Trev.) Meisn by the method of subrenal capsule(SRC) assay. A total of 12 lung cancers had been tested in which 60 xenografts were implanted. With a mean growth of  greater than 5 OMU (1 OMU=0.02 mm) in the control animal an assay was considered evaluable. This study provided an evaluable assay rate of 83.3%. Taking  less than xe2x88x9210 OMU of tumor xenograft size as the standard of responsiveness extracts E and CD1 offered response rates of 40% and 20% respectively. Squamous cell carcinoma was more sensitive to E than other types of lung cancer. These findings were similar to the clinical effects of CD1 phase-I study. The study also confirmed that extracts E and CD1 had no toxicity or side effects in mice.(10)
Liang X Z et al. described using TLC scanning to determine procyanidin B-2 in Fagopyrum dibotrys""s rhizome in an article published in China in 1990. The authors reported that procyanidin B-2 in Fagopyrum cymosum was determined by TLC scanning, taking high efficient silica gel 60F254 as thin-layer absorbent, toluene-methylformaic-methanol-formic acid (1:2:0.2:0.1) as developing agent, Rf=0.21, scanned at 280 nm with CAMAG 76510 TLC scanner. The sample was prepared by leaching for 62 hours in methanol. The average recovery was 98%. Standard deviation was 0.0092, coefficient of variation is 1.8%(n=10).(11)
Liang, M D et al. published an article in 1991 on extra corporeal anti-cancer function of Fagopyrum cymosum rootin. The authors pointed out that Fagopyrum cymosum rootin (FCR) is a new anti-cancer medicine screening from Fagopyrum cymosum roots, with the method of extra corporeal culture of human cancer cell. The active constituent is large-moleculared condensed tannin D. At the concentration of 125 xcexcg/ml, its inhibition ratio is 84.5% for SGC, 78.9% for Hela and 100% for KB, caused by the impairment of the cancer cell""s membrane, RNA, DNA metabolization and cell nucleus cleavage.(12)
In the same year, Ma MF et al. reported the results of mutagenicitic and teratogenicitic tests of Fagopyrum cymosum (Trev.) Meisn. The results showed that no positive mutation was induced in four Ames standard bacterial strains (xc2x1S9), using seven different doses of Fagopyrum cymosum (Trev.) Meisn varying from 1-5,000 xcexcg/plate. Antimutational effect was exhibited against reversional mutation of TA98 and TA 100 strains induced by daunorubicin and methyl methane sulphonate. No increase in micronucleus frequency in bone marrow polychromatophil erythroblast of NIH mice, no teratogenicity in Chinese hamster oocyte chromosomes (xc2x1S9), no adverse effect on reproductive capacity of NIH mice and development of mouse fetuses, nor teratogenicity of appearance, skeleton and internal organs of mouse fetuses were observed.(13)
Gao Z. et al. described in 1993 the effects of Fagopyrum cymosum root extract on proliferation of four human tumor cells in vitro in comparison with 5-FU. The researchers found that the proliferation of four human tumor cells were markedly inhibited by the extract and the inhibition rates were positively proportional to concentration. At the concentration of 12.5 micrograms/ml, the inhibition rates were 98.7% for GLC, 82.1% for KB, 65.4% for SGC and 53.8% for Hela cells.(14)
Zhang W J et al. discussed phenolic constituents from Fagopyrum dibotrys in 1994. The researchers reported that six phenolic constituents had been separated from the alcohol extracted powder of the dry rhizome of Fagopyrum cymosum (Trev.) Meisn. By polarimetry, HNMR, 13C-NMR and FAB-MS, they were identified as: 3,4-dihydroxybenzoic acid, gallic acid, (xe2x88x92) epicatechin, (xe2x88x92) epicatechin-3-O-gallate, procyanidin B-2 and procyanidin C-2.(15)
In the same year Meng F H et al. published an article on anticancer effect of cymose buckwheat roots on human tumor cells cultured in vitro. The researchers reported that anticancer effect of cymose buckwheat roots on various human cancer cells cultured in vitro had been studied using direct killing method, colony inhibition test and 3H-TdR incorporation test. The results showed that the drug at the concentration of 1 gxc2x7Lxe2x88x921 had a killing rate of over one logasithmic killing against a number of human cancer cells, and when the concentration was lowered to 0.125 gxc2x7Lxe2x88x921, its killing rate could still reach 74.3-92.1%, approximating one logasithmic killing. The extract from the roots of cymose buckwheat had significant anticancer effect, showing a colony inhibiting rate of 100% against several cancer cells when its concentration was 0.1 or 0.05 gxc2x7Lxe2x88x921, and that of 75.1-89.2% at the concentration of 0.0125 gxc2x7Lxe2x88x921. 3H-TdR labeling revealed that the drug could significantly inhibit nucleic acid metabolism in cancer cells, the inhibitory effect being close to that of positive control, fluorouracil, at the same concentration.(16)
In another article published in the same year, Meng F H et al. reported anticancer effect of Jin E in vitro. Jin E is a compound of tannin extracted from cymose buckwheat. In the studies on anticancer effect of Jin E, direct killing method, colony inhibition test and 3H-TdR radioautography were applied. Results of the studies showed that the 50% inhibiting concentrations of Jin E against GLC, Hela, SGC and KB cells were 67.8, 73.1, 79.9 and 83.0 xcexcg/ml respectively, with a definite concentration and time-response relationship. Jin E at the concentrations of 100 and 50 xcexcg/ml could completely inhibit colony formation of several human cancer cells, and Jin E at the concentration of 25 xcexcg/ml had a colony inhibiting rate as high as 95% against four kinds of human cancer cells; the best anticancer activity was observed in Jin ED and alcohol soluble portion. Jin E at the high concentration (100 xcexcg/ml) had an inhibiting rate of 87.9% against intracellular nucleic acid synthesis, the difference being very significant (P less than 0.01), compared with blank control group. It is concluded that Jin E has significant inhibitory effect on growth of multiple human cancer cells in vitro.(17)
Peng Yong et al. reported the research and development of Fagopyrum dibotrys in a paper published in China in 1996. The paper reviewed new developments in Fagopyrum dibotrys studies in the areas of the materia medica, the plant, the chemistry, the pharmacology, the clinic applications and the preparation of the medicine. The authors also provided suggestions on further exploitation and utilization of Fagopyrum dibotrys.(18)